Organic Carbon Controls Mercury Distribution and Storage in the Surface Soils of the Water-Level-Fluctuation Zone in the Three Gorges Reservoir Region, China

The particular condition of the water-level-fluctuation zone (WLFZ) in the Three Gorges Reservoir (TGR), the largest hydroelectric reservoir in China, raises great concerns about mercury (Hg) contamination and ecological risk. In addition, previous research found that soil organic carbon (SOC) plays an essential role in controlling Hg distribution and speciation. However, there is minimal information on the Hg storage distribution and their relationships with SOC in the WLFZ in TGR. This study investigated Hg distribution, storage, and their relationships with SOC in the surface soils in WLFZ. The results showed that the total Hg (THg) content in the surface soils ranged from 18.40 to 218.50 ng g−1, with an average value of 78.17 ± 41.92 ng g−1. About 89% of samples had THg content above the background value in Chongqing, showing specific enrichment of Hg in WLFZ due to contamination in the TGR. The surface soils have low SOC, with an average value of 8.10 ± 3.90 g kg−1. Moreover, THg content showed consistent distribution with the SOC in WLFZ, with a significantly positive correlation (R = 0.52, p < 0.01, n = 242). THg storage (201.82 ± 103.46 g ha−1) in the surface soils was also significantly positively correlated with the SOC storage (R = 0.47, p < 0.01, n = 242). The reduced SOC sequestration, due to the periodical alternative “flooding–draining” and frequent reclamation and utilization of WLFZ, decreased the Hg adsorption in soil. Those might result in the re-release of Hg into waters when WLFZ is flooded. Therefore, more attention should be directed towards Hg cycling and the consequent environmental risks in the TGR region.


Introduction
Mercury (Hg) is one of the most hazardous pollutants with potent toxicity [1,2]. Hg may be present in three chemical forms: elemental mercury (Hg 0 ), divalent inorganic mercury (Hg II ), and methylmercury (MeHg) [3]. It has received worldwide attention for decades due to its high toxicity, prevalent existence, and bioaccumulation through the food chain of its methylation product: MeHg [4,5]. The biogeochemical cycles of Hg can be influenced by soil organic carbon (SOC) through several aspects, including ligand binding [6] and redox processes [7]. Firstly, research shows that the adsorption/desorption of Hg species with sediment particles, which is mediated by the partitioning of associated organic ligands, is a primary driving force for the distribution of MeHg and Hg II [8]. Based on studies on sediments, and other high-SOC sites, it is generally accepted that SOCrich soils show a greater capacity to immobilize Hg due to their binding and adsorption capacities [9][10][11]. Secondly, reducing Hg II to Hg 0 via SOC accounts for an important part of legacy Hg recycling back into the atmosphere [12][13][14]. Thus, untangling the relationships Int. J. Environ. Res. Public Health 2023, 20, 3681 3 of 13 area in this study is located in the Chongqing section of the TGR region (106 • 50 -110 • 50 E, 29 • 16 -31 • 25 N) between the Jiangjin District and Wushan County. The climate of the TGR area is subtropical monsoon, with an annual average temperature of 17.9 • C, sunshine duration of approximately 1630 h, average frost-free period of approximately 260 d, and annual precipitation of 1000-1800 mm [32]. Contrary to the natural wetting-drying aquatic system such as lakes and rivers, the anti-seasonal water level management controls the water level in TGR areas, changing from the 145 m a.s.l. (summer, called dry period) to 175 m a.s.l. (winter, called wet period). As a result of such non-seasonal fluctuations in the water-level, WLFZ is formed within a vertical height of 30 m (approximately a total area 350 km 2 ) [38]. The main soil type in this area is purple soil and barren and agricultural lands (planted with corns by the local farmers from March to August) are the major land types in this area. From 2012-2013, during the two dry periods (i.e., from May to August) of each year, two sampling campaigns were conducted. In total, 15 sites were selected for soil sampling. The sampling sites cover Wushan, Fengjie, Yunyang, Kaixian, Wanzhou, Shizhu, Zhongxian, Fengdu, Feiling, Changshou, Banan, Nan'an, Jiangbei, Yubei, and Jiangjin ( Figure 1), which are the administrative districts and counties in Chongqing city. Samples were harvested from the surface soil layer (0-20 cm). According to the sampling guide [39], one sampling plot was set for at least 10 m × 10 m. The five soil cores were obtained from the middle and four corners of each plot, which were further combined to form a composite sample. Overall, (n = 242) soil samples were collected in total. All soil samples were transported to the laboratory on ice in polyethylene plastic bags.

Study Area and Sample Collection
The TGR, the largest hydroelectric reservoir in China, has a total area of flood landscapes of 630 km 2 . Among them, 350 km 2 was a seasonally flooded WLFZ which is heavily influenced by human activities along the TGR. The TGR region is located in the 600 km section of the Yangtze River between the cities of Chongqing and Yichang. The area in this study is located in the Chongqing section of the TGR region (106°50′-110°50′ E, 29°16′-31°25′ N) between the Jiangjin District and Wushan County. The climate of the TGR area is subtropical monsoon, with an annual average temperature of 17.9 °C, sunshine duration of approximately 1630 h, average frost-free period of approximately 260 d, and annual precipitation of 1000-1800 mm [32]. Contrary to the natural wetting-drying aquatic system such as lakes and rivers, the anti-seasonal water level management controls the water level in TGR areas, changing from the 145 m a.s.l. (summer, called dry period) to 175 m a.s.l. (winter, called wet period). As a result of such non-seasonal fluctuations in the waterlevel, WLFZ is formed within a vertical height of 30 m (approximately a total area 350 km 2 ) [38]. The main soil type in this area is purple soil and barren and agricultural lands (planted with corns by the local farmers from March to August) are the major land types in this area. From 2012-2013, during the two dry periods (i.e., from May to August) of each year, two sampling campaigns were conducted. In total, 15 sites were selected for soil sampling. The sampling sites cover Wushan, Fengjie, Yunyang, Kaixian, Wanzhou, Shizhu, Zhongxian, Fengdu, Feiling, Changshou, Banan, Nan'an, Jiangbei, Yubei, and Jiangjin ( Figure 1), which are the administrative districts and counties in Chongqing city. Samples were harvested from the surface soil layer (0-20 cm). According to the sampling guide [39], one sampling plot was set for at least 10 m × 10 m. The five soil cores were obtained from the middle and four corners of each plot, which were further combined to form a composite sample. Overall, (n = 242) soil samples were collected in total. All soil samples were transported to the laboratory on ice in polyethylene plastic bags.

Analysis Method
The soil pH was measured by making soil slurry in a soil-water ratio of 1:2.5 (w/v). The mixture is stirred for 3 min and then filtered under gravity for a 30 min period. The filtrate's pH represents the soil's pH, and it is measured with a portable pH meter (ST300, OHRUS ® , Cole-Parmer, Wertheim, Germany) [40]. The amorphous Fe oxide (Feo) contents of the bulk soil samples were determined by ammonium oxalate buffer solution [41]. The

Analysis Method
The soil pH was measured by making soil slurry in a soil-water ratio of 1:2.5 (w/v). The mixture is stirred for 3 min and then filtered under gravity for a 30 min period. The filtrate's pH represents the soil's pH, and it is measured with a portable pH meter (ST300, OHRUS ® , Cole-Parmer, Wertheim, Germany) [40]. The amorphous Fe oxide (Fe o ) contents of the bulk soil samples were determined by ammonium oxalate buffer solution [41]. The soil cation exchange capacity (CEC) was measured using the sodium acetate method [42]. The physical and chemical properties of soil were presented in Table 1. Hg in soil was determined via thermal decomposition atomic absorption spectrometry after gold amalgamation, using DMA-80 (Milestone, Italy). Two method blanks, three certified reference materials (CRMs), and 10% replicate samples were accompanied in each sample batch (up to 30 samples) for quality assurance (QA) and quality control (QC) of sample detection. The method detection limit (MDL) of Hg in soil was 0.009 ng g −1 .
The method blanks were lower than the detection limits in all cases. The SD of sample duplicates ranged from 0.28-10.5%. The recovery rate for CRMs in soil (GBW07406) ranged from 86-107%. A potassium dichromate external heating method measured the SOC. The soil bulk density was measured using the cutting-ring method [43].
The Hg storage in the soil of WLFZ in the TGR area is calculated as: where S Hg is the total Hg storage in the soil (g ha −1 ); C i is the Hg content in the soil (ng g −1 ); ρ i is the soil bulk density (g cm 3 ), and h is the soil thickness (cm). The SOC storage in the soil is calculated as: where SOC D is the organic carbon storage in the soil (kg m −2 ); SOC i is the organic carbon content in soil (g kg −1 ); ρ i is the soil bulk density (g cm −3 ); h is the soil thickness (cm); and δ is the proportion of particles with a diameter larger than 2 mm.

Statistical Analysis
Data processing and analysis were conducted using SPSS 26.0 (IBM, Armonk, NY, USA) and the "ggplot2" package in the R version 4.2.2 [44]. Partial least square path modeling (PLS-PM) was used to confirm the correlations of soil chemical properties (pH, Fe o , CEC, SOC) with Hg distribution and storage. The partial least square (PLS) analysis was developed in 1960 to compensate for the limitations of multivariate normality and large sample sizes in the analysis using the existing linear structural relationships [45]. In particular, it has the ad-vantage of allowing analysis even when only a small number of samples are available. Path modeling (PM) is used to evaluate the validity and reliability of measurements and analyze the causal relationships among tested variables.

Distribution of Hg in the Surface Soils
The Hg contents in the surface soils of WLFZ in TGR ranged from 18.40 to 218.5 ng g −1 , with a mean value of 78.17 ± 41.92 ng g −1 . They varied significantly, with a variation coefficient of 53.63% (Table 2). Moreover, 89% of samples exceeded the background value in Chongqing (37.00 ng g −1 ) [46]. The average Hg content varied significantly and ranged from 38.9 to 125.5 ng g −1 , which was 1.1-3.4 times higher than the background value in Chongqing [46]. The most considerable Hg content reached 218.5 ng g −1 (Figure 2a), 5.9 times the background value, indicating the particular Hg contamination in soils in the study area. The most serious contamination was found in Jiangbei, Fuling, and Nan'an, with the Hg contents in all the samplings sites above the background value. In contrast, although low content was found in Fengdu and Shizhu, 75% of samples had values above the background values. "N" means number of samples; "Min" means the minimum value; "Max" means the maximum value; "SD" means the standard deviation; "CV" means variable coefficient.

Distribution of the Organic Carbon in the Surface Soil
The SOC content ranged from 2.28 to 23.79 g kg −1 ( Table 3). The highest SOC content (12.20 ± 4.64 g kg −1 ) was found in Fengjie, and the lowest values (about 5.5 g kg −1 ) were found in Fengdu, Yunyang (Figure 2b). The SOC content in WLFZ in TGR was relatively low compared to that in other wetland areas in China [23,47]. The SOC (0-15 cm) in the Wanjiang wetland, located in the middle and lower reaches of the Yangtze River, was about 11.30-27.83 g kg −1 , with a mean value of 17.00 ± 1.50 g kg −1 [23]. The SOC of the Dongting Lake wetland was above 40 g kg −1 [47].
Hg storages in the soil in WLFZ in the TGR varied significantly, with a range value of 50.76-549.58 g ha −1 and a variation coefficient of 51.36% (Figure 2c). The average Hg storage was 201.83 ± 103.46 g ha −1 in the study area. The Hg storage showed different distribution from that of Hg content in a different area, which might be due to the different bulk densities. The highest Hg storage (318.43 ± 77.27 g ha −1 ) was found in Nan'an, and the lowest (102.82 g ha −1 ) was in Shizhu.

Distribution of the Organic Carbon in the Surface Soil
The SOC content ranged from 2.28 to 23.79 g kg −1 ( Table 3). The highest SOC content (12.20 ± 4.64 g kg −1 ) was found in Fengjie, and the lowest values (about 5.5 g kg −1 ) were found in Fengdu, Yunyang (Figure 2b). The SOC content in WLFZ in TGR was relatively low compared to that in other wetland areas in China [23,47]. The SOC (0-15 cm) in the Wanjiang wetland, located in the middle and lower reaches of the Yangtze River, was about 11.30-27.83 g kg −1 , with a mean value of 17.00 ± 1.50 g kg −1 [23]. The SOC of the Dongting Lake wetland was above 40 g kg −1 [47].   "N" means number of samples; "Min" means the minimum value; "Max" means the maximum value; "SD" means the standard deviation; "CV" means variable coefficient.

Hg Storage in the Surface Soil 3.4. The SOC Storage in the Surface Soil
The estimated SOC storage of WLFZ in TGR was shown in Figure 2d. In the study area, the SOC storage ranged from 0.63 to 5.28 kg m −2 , with an average value of 2.09 ± 0.95 kg m −2 . The SOC storages varied with different districts/counties, with a variation coefficient of 14.56~48.26%. The highest SOC storage was found in Fengjie (3.06 ± 0.99 kg m −2 ), while the lowest was in Yunyang (1.41 ± 0.43 kg m −2 ).

The Physical and Chemical Properties of the Surface Soil
The physical and chemical properties of WLFZ in TGR was shown in Table 1. In the study area, the pH ranged from 4.73 to 8.54, with an average value of 7.54 ± 0.79. CEC and Fe o in the soil in WLFZ in the TGR varied significantly, with a range value of 3.77-42.27 cmol kg −1 and 1067.50-6923.26 mg kg −1 , respectively. The highest CEC (21.06 ± 9.95 cmol kg −1 ) was found in Fuling, and the lowest (10.81 ± 7.09 cmol kg −1 ) was found in Banan.

Distribution and Storage of Hg in the Surface Soil
Chongqing is a fast-developing industrial region in southwest China. Despite the control and management regulation of potentially toxic elements contamination in recent years, potential toxic elements, including Hg, might accumulate in the soils through atmospheric deposition due to the extensive production and emissions that occurred in the past. Those Hg might enter into the soils in TGR via rain wash and surface runoff during the draining period. The Hg storage here was much lower than previous measurements (102.9 ± 9.8 mg m −2 ) in the soil (0-40 cm) of Chongqing [48]. This might be due to the different sampling depths (0-20 cm), in which the Hg in surface soil could be reduced by washing off with waters and re-released into waters via the soil/water surface during the flooded period. Furthermore, these factors reduced the Hg storage but increased the environmental risk. In this study, the coefficient of variations (CV) of Hg contents was 53.63%, indicating large variations in spatial distributions for Hg. However, the correlation analysis found the Hg storage significantly positively correlated with the SOC in the soils in the WLFZ, with a Pearson correlation coefficient of 0.47 (p < 0.01, n = 242) (Figure 3a). This suggests the SOC storage in the soil greatly affected Hg storage regardless of the spatial variations of Hg distributions in WLFZ.

Distribution and Storage of SOC in the Surface Soils
Like other ecosystems, such as wetlands, WLFZ is substantially influenced by water levels. Thus, SOC turnover and accumulation due to oxic-anoxic degradation and plant debris inputs are active in WLFZ. The SOC content in WLFZ in TGR was relatively low compared to that in other wetland areas in China. The WLFZ in TGR represents a transition between aquatic and terrestrial systems with low soil gleization, vegetation/organisms, and the rapid decomposition of animals and plant residues. Under the periodical alternative wetting-drying condition in TGR, the hydrological characteristics, including water level fluctuation, velocity, and flow, varied significantly, resulting in a significant loss of soil as well as the residues of animals and plants, which decreased SOC content and carbon sequestration.
Previous studies found that the wetlands showed soil carbon sequestration and carbon sink [49], with the higher SOC storage in Wanjiang and Dongting lake wetlands discussed above. With wetland reclamation, the SOC content in the soil might decrease due to the reduced input of SOC and enhanced decomposition [23]. Here, the SOC storage was low compared to that in the two wetlands mentioned above. There are two potential explanations for this. One is that the majority of the WLFZ is utilized as agricultural soil by local peoples, resulting in a reduction in the SOC storage. A previous study has reported that conversion of non-cultivated land for agricultural purposes has substantially reduced global SOC stocks in upper soil layers, and the results are consistent [50]. The other possible explanation is that, as a new reservoir, WLFZ has limited soil carbon sequestration. Similar research has shown that reservoirs emit more carbon than they bury, challenging the current understanding that reservoirs are net carbon sinks [51].

Distribution and Storage of SOC in the Surface Soils
Like other ecosystems, such as wetlands, WLFZ is substantially influenced by water levels. Thus, SOC turnover and accumulation due to oxic-anoxic degradation and plant debris inputs are active in WLFZ. The SOC content in WLFZ in TGR was relatively low compared to that in other wetland areas in China. The WLFZ in TGR represents a transition between aquatic and terrestrial systems with low soil gleization, vegetation/organisms, and the rapid decomposition of animals and plant residues. Under the periodical alternative wetting-drying condition in TGR, the hydrological characteristics, including water level fluctuation, velocity, and flow, varied significantly, resulting in a significant loss of soil as well as the residues of animals and plants, which decreased SOC content and carbon sequestration.

Correlation between Hg Distribution, Storage, and SOC
The distribution of the Hg storage significantly positively correlated with the SOC storage (R = 0.47, p < 0.01, n = 242) (Figure 3a). One study found that Hg storage was significantly positively correlated with SOC and nitrogen in the soil in four forests in the Sierra Nevada [52]. This might be explained by the increased soil adsorption capacity, due to great surface litter in the forests, which enhanced the accumulation of atmospheric Hg deposited into the soil with the leaves. However, the surface litter was very limited in the WLFZ in the TGR under the alternative wetting-drying condition, resulting in low SOC content. Additionally, the great population and agricultural/industrial activities along the WLFZ could affect SOC storage. Therefore, the WLFZ was heavily influenced by humaninduced disturbances and the periodical alternative wetting-drying condition. Those greatly affect carbon sequestration and Hg storage in WLFZ. The carbon sequestration and SOC storage in WLFZ were reduced due to the short formation time, frequent utilization, and reclamation. These factors might decrease the Hg absorption in the soil in WLFZ, resulting in the potential re-release of Hg into waters during the flooding period.
The Hg content in the soil of WLFZ in TGR was significantly positively correlated with that of SOC, with a Pearson correlation coefficient of 0.52 (p < 0.01, n = 242) (Figure 3b). The highest Hg content (218.5 ng g −1 ) and SOC (23.79 g kg −1 ) were found in Jiangbei, while the lowest Hg content (18.4 ng g −1 ) and SOC (2.34 g kg −1 ) was found in Fengdu. In addition, the SOC greatly affects the degradation of SOC and the soil's CEC [53]. In this study, both the CEC and SOC significantly correlated with the Hg content, with correlation coefficients of 0.52 and 0.19 (p < 0.01, n = 242) (Figure 3b,c), respectively, implying that the SOC could influence Hg behavior in the soil as a function of the CEC. In addition, the Fe o content in soils plays an important role in SOC stabilization, and the formation of organo-mineral complexes has been recognized as a critical mechanism [54]. Our results revealed that the SOC content in WLFZ in TGR significantly positively correlated with the Fe o (R = 0.208, p = 0.001, n = 242) (Figure 3d). Additionally, the Fe o and the Hg content showed a significantly positive correlation (R = 0.261, p < 0.001, n = 242) (Figure 3e), indicating Fe o was a critical driving factor that significantly influenced the Hg and SOC storage in the soils of TGR. Furthermore, regarding the coupling between Fe o and SOC for the SOC stabilization potential, the results (Figure 3a) indicate that in TGR, a higher stabilization degree of SOC shows greater capacities for holding Hg storage. This suggests that enhancing SOC stabilization might be a suitable strategy for offsetting the greenhouse emission feedback to global warming. Meanwhile, this methods helps to stabilize more Hg inputs from atmospheric deposits, resulting in less Hg losses from soils into nearby aquatic systems.

Partial Least Squares Path Modeling (PLS-PM) Analysis
Multivariate statistics analysis achieved via PLS-PM was selected to evaluate the influencing roles and relative contributions of pH, Fe o , CEC, and SOC on Hg distribution and storage. SOC includes SOC distribution and storage, and Hg is a collection of Hg distribution and storage. The standardized direct effects of pH, CEC, Fe o , and SOC on the Hg distribution and storage are shown in Figure 4. The goodness of fit (GoF) was 0.42, and SOC had the greatest direct positive effects (0.46) on Hg distribution and storage. This suggests that SOC distribution and storage determined the Hg distribution and storage, further supporting our correlation analysis results between Hg and SOC. Studies have reported a close link between SOC and Hg [55][56][57][58]. This study further demonstrates that SOC distribution and storage roles are predominant in controlling Hg distribution and storage.  Figure 4. The goodness of fit (GoF) was 0.42, and SOC had the greatest direct positive effects (0.46) on Hg distribution and storage. This suggests that SOC distribution and storage determined the Hg distribution and storage, further supporting our correlation analysis results between Hg and SOC. Studies have reported a close link between SOC and Hg [55][56][57][58]. This study further demonstrates that SOC distribution and storage roles are predominant in controlling Hg distribution and storage.

Conclusions
Although we have explored mercury (Hg) biogeochemistry in the water-level fluctuation zone (WLFZ) of the Three Gorges Reservoir (TGR) areas to some degree during the past few years, this study provides a whole picture of Hg distribution and its relationship

Conclusions
Although we have explored mercury (Hg) biogeochemistry in the water-level fluctuation zone (WLFZ) of the Three Gorges Reservoir (TGR) areas to some degree during the past few years, this study provides a whole picture of Hg distribution and its relationship with environmental factors, especially soil organic carbon (SOC). We found that the Hg contents varied greatly in the surface soils of WLFZ in the TGR areas, and the average Hg content was significantly greater than the background value in Chongqing. Additionally, the spatial distributions of Hg contents and storage were linked with SOC content and amorphous Fe oxides (Fe o ) in soils within significant correlations. These findings validated our expectation that SOC is crucial in controlling Hg storage in soils. Regarding the association between Fe oxides and SOC in regulating SOC stabilization, our results support SOC stabilization's potential role in Hg storage in these areas. Based on the these extensive insights, destabilization of SOC resulted in harsher feedback of the soil carbon pool in relation to climate change, which also could elevate the releases and emissions of Hg that was stored previously. As a result, increases in environmental risks induced by Hg elevations could be expected in the context of the changing climate. Thus, policies and management for stabilizing SOC in the TGR areas will mitigate climate change feedback. It could also be beneficial to control Hg pools as sinks rather than sources. In the future, great attention and further study will be needed to examine the coupling between the carbon and mercury cycles in WLFZ in TGR areas.
Author Contributions: S.Z.: methodology, investigation, data curation and analysis, and writingoriginal draft preparation; C.Y.: data curation and analysis, discussion, and writing-reviewing and editing; H.C. and Y.W.: discussion, and writing-reviewing; J.L.: methodology, investigation, and writing-original draft preparation; R.Z. and Y.Y.: methodology, data curation and analysis, discussion, and writing-reviewing and editing; D.W.: conceptualization, discussion, and writingreviewing and editing; C.Z.: project supervision; conceptualization, methodology, data curation and analysis, writing-original draft preparation, writing-reviewing and editing. All authors have read and agreed to the published version of the manuscript.

Institutional Review Board Statement:
We have not included anything regarding ethical issues in this experiment, such as human or animal models, so this is not applicable.

Informed Consent Statement: Not applicable.
Data Availability Statement: Data are available from the corresponding author upon reasonable request.

Conflicts of Interest:
The authors declare no conflict of interest.

WLFZ
The water-level fluctuation zone TGR The